Over recent years cell culture has become a core technology in the life sciences. Cell culture is described in ‘Basic Cell Culture’ Oxford University Press (2002) Ed. J. M. Davis; and ‘Animal Cell Culture’ Oxford University Press (2000) Ed. John R. W. Masters; both of which are incorporated herein in their entirety by reference. Cell culture provides the basis for studying cellular processes such as the viability, phenotype, genotype, proliferation and differentiation of cells, and the formation of biological molecules, intermediates and products. It has also provided the means to study the regulation of these processes, from the genetic level-whether in isolation or in whole transgenic animals-down to the level of individual protein molecules.
Notwithstanding its enormous contribution to the current state of biology, in many respects cell culture remains a developing discipline, albeit an unusually exciting science ultimately offering the possibility of genetic therapy and tissue engineering. An important goal of cell culture is to be able to grow a wide variety of cells in vitro. The list of different cell types that can be grown in culture is extensive (see American Type Culture collection, http://www.atcc.org; European Collection of Cell Cultures, http://www.ecacc.org.uk; Deutsche Sammlung von Mikroorganismen and Zellkulturen GmbH, http://www.dsmz.de), includes representatives of most cell types, and is continually increasing as more and more culture conditions are discovered.
Despite the steady progress in the field, until recently the method of determining suitable culture conditions for new cell types has remained totally empirical: growth conditions are almost always discovered by trial and error. The choice of starting point will often be based on what was previously used by others for similar cells, or even what is currently being used in the laboratory for different cells. Many times these will simply be completely inadequate, and a process of trial and error must begin anew. Even when new culture conditions are successful, it is worthwhile remembering that adaptations of previous protocols will have introduced a historical bias to the experiment. For instance, much of the early tissue culture experiments made extensive use of fibroblasts, and to this date most standard cell culture conditions favour growth of cells derived from the mesoderm (fibroblasts, endothelium, myoblasts). The development of selective growth media for epithelial and other cell types based on these conditions was a challenge. For some of these cell types it is now known that serum—a normal component of many culture media for mesodermal cells-actually inhibits growth.
Applicants have previously described methods for developing suitable culture conditions, which allow for the viability, proliferation or growth, and retention of the phenotype of particular cell types. Some common problems, which are still encountered in cell culture, are the limited lifespan of primary cell lines, the change of characteristics of cell lines with passage, and their transformation accompanied by loss of interesting cellular characteristics. These effects severely limit the utility of cultured cells for use in experiments or assays.
Improved techniques for culturing cells and methods for discovering and implementing techniques for regulation of cellular processes such as growth, differentiation, metabolic activity, and phenotypic expression are presented in Applicants' international application WO 2004/031369. According to the procedures described therein, “units” of cells, which comprise one or more cells cultured, for example, on a bead, are subjected to different growth conditions in a combinatorial split-pool procedure, which involves repeated splitting and re-pooling of cell cultures, to expose different cell units therein to different culture conditions.
When handling large numbers of cell units, their identity and/or cell culture history (for example, the chronology and the exact nature of a series of culture conditions that any one group or unit may have been exposed to) can become confused. WO2004/031369 describes improved methods for determining the identity and/or cell culture history of cell units.
In WO2007/063316, Applicants describe methods for determining the activity of agents which act on a cell, using the split-pool procedure.
In WO2007/023297, Applicants describe further improved methods for tagging cells in split-pool cell culture experiments, better to determine which reagents and nutrients a cell has been exposed to in achieving a particular state.
The performance of split-pool cell culture experiments, however, is manually laborious and time-consuming, requiring iterative transfer of cell units between culture vessels and optionally between filtering devices and culture vessels. The transfer of cell cultures in between individual vessels for filtering, pooling, splitting and subsequent rounds of the same moreover potentially exposes the cultures to infectious agents and other sources of contamination. There is therefore a need for a cell culture vessel in which split-pool culture can be conducted easily, without the need to transfer the culture continuously between vessels.
Citation or identification of any document in this application is not an admission that such document is available as prior art to the present invention.